In 2001, our group began studies to determine the genetics causes of bone fragility. We chose to study rats, rather than mice, because the laboratory rat has proven to be an excellent model for human bone disorders, like osteoporosis. Because rats are larger than mice, it is easier to measure bone structure and strength at the femoral neck, which is a skeletal site of primary focus. Our first study involved a cross between Fischer 344 (F344) and Lewis (LEW) rats. Our second study focusing on Copenhagen 2331 (COP) and Dark Agouti (DA) rats began in 2003 and is the subject of this competitive renewal application. We have successfully mapped quantitative trait loci (QTLs) for femoral neck and femoral midshaft phenotypes, the primary and secondary goals of the project. Our work in the past five years has provided a clear direction for our proposed research in the next five years: we will identify genes within QTLs that affect bone biology. This project has three Aims. First, we will identify causative genes within our two QTLs with largest effect size: Chromosome (Chr) 4 for F344 and LEW rats (13% effect) and Chr 1 for COP and DA rats (14% effect). We plan to generate congenic rat models and conduct gene expression profiling followed by functional studies in vivo and using cultured osteoblasts and osteoclasts to identify genes that directly affect bone biology. Second, we will work with a large, world-wide research consortium to conduct a genome wide association study (GWAS) in heterogeneous stock (HS) rats. The GWAS led by Jonathan Flint will provide phenotypes and genotypes for over 2000 rats. Phenotypes will include: behavioral, metabolic, hematological, hemodynamic, immunological and skeletal (our contribution). Each rat will be genotyped for about 20,000 single nucleotide polymorphisms (SNPs). The results will allow us to identify new QTLs that affect bone traits and to reduce the number of candidate genes at each QTL from several dozen to a mere handful. Finally, we will identify gene expression QTLs (eQTLs) using a genome screen to map transcript abundance in 2nd filial (F2) rats derived from COP and DA progenitors. Variations in expression of individual genes will be mapped to locations on the genome to produce eQTLs. eQTL mapping allows one to identify cis-eQTLs for which expression levels can be correlated with a chosen bone phenotype to identify and prioritize genes most likely to alter bone biology. Also, we will identify trans- genes controlled by a given QTL and with this information we will construct networks of genes that underlie complex bone traits. Another unique feature of eQTL mapping is its ability to identify allele-specific regulation of gene expression on a genome-wide scale. Completion of these Aims will accelerate our progress toward identifying genes that affect bone fragility. PUBLIC HEALTH RELEVANCE: Our study will identify genes that cause bone fragility. We will use several inbred strains of rats in our experiments and apply modern genetics techniques like gene expression microarrays and single nucleotide polymorphism genotyping. Our goal is to find genetic causes for bone weakening conditions like osteoporosis so better treatments can be developed.